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Detection of 1-ß-D-Arabinofuranosylcytosine Incorporation into DNA in Vivo¹

Laboratory of Clinical Pharmacology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115

The incorporation of (1-ß-d-arabinofuranosylcytosine (ara-C) into the DNA of leukemic cells is highly correlated with cytotoxicity in vitro. However, the measurement of ara-C incorporation into leukemic cell DNA in vivo during ara-C therapy has been limited by the lack of a suitably sensitive method. A quantitative assay procedure has therefore been developed to determine incorporation of unlabeled ara-C into DNA. This method involves DNA isolation from patient myeloblasts, enzymatic digestion of the DNA, high pressure liquid chromatography separation of the nucleosides, and determination of ara-C in the eluate fractions by radioimmunoassay. Using this approach, incorporation of unlabeled ara-C into DNA of HL-60 cells is log linear over concentrations of 1 to 100 µM ara-C. Furthermore, the extent of ara-C incorporation into DNA as determined by this method correlates significantly with measurements of [³H]ara-C (DNA) formation under similar conditions. This approach has also been applied to clinical samples. Myeloblasts from 6 patients receiving high-dose continuous-infusion ara-C therapy incorporated 0.00–0.36 pmol ara-C/µg DNA during 24 h of therapy. These findings thus suggest that this method can be used to monitor the in vivo incorporation of ara-C into leukemic cell DNA.

¹ Supported by USPHS Grants CA 29431 and K08CA00994 (D. S.) awarded by the National Cancer Institute, Department of Health and Human Services, and by a Burroughs Wellcome Award in Clinical Pharmacology (D. K.).

² To whom requests for reprints should be addressed, at Laboratory of Clinical Pharmacology, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115.

Received 4/16/87. Revised 9/21/87. Accepted 9/22/87.